目的研究不同剂量X射线诱导乳腺癌细胞株MCF-7自噬发生过程中beclin1表达的变化和作用。方法采用实时荧光定量PCR方法检测自噬相关基因MAP1LC3B表达,RT—PCR方法检测beclin1基因变化,X射线Western blot方法检测beclin 1蛋白表达。结果MAP1LC3B mRNA量效研究表明,8、16和32h量效研究均发现表达上调,8、16h在12Gy,32h在8Gy达到表达峰值,总体表现为表达升高(F=13.831、10.996、17.019,P〈0.05);时程研究表明,2Gy时程研究发现MAP1LC3B于16h升高达至峰值(F=16.284,P〈0.01),8Gy时程研究中32h升高达至峰值(F=9.030,P〈0.05),12Gy时程研究中16h升高达至峰值(F=20.315,P〈0.05)。beclin 1 mRNA量效关系表明,照射后4h beclin 1在2、4和8Gy阶段呈剂量依赖增加趋势;8Gy时间效应关系中发现照射后beclin 1 mRNA的变化呈时间依赖性增加。16h蛋白量效关系表明,照射后beclin 1表达增加;2Gy时效关系表明照射后蛋白表达增加。结论X射线照射能够诱导乳腺癌细胞株MCF-7发生自噬,beclin 1基因上调表达的变化提示其发挥重要作用。
Objective To investigate the changes of beclin 1 expression in radiation-induced autophagy in breast cancer cell line MCF-7. Methods Real-time RT-PCR was used to detect the expression of autophagy-related genes MAP1LC3B. Reverse transeription-PCR was used to detect the expression of beclin 1. Western blot was used to analyze the protein expression of beclin 1 . Results Dose-response ofMAP1LC3B mRNA studies showed that MAP1LC3B mRNA increased in 8, 16 and 32 h dose-effect studies, up to peak at 12 Gy in 8 h study(F = 13. 831 ,P 〈 0. 05) , 12 Gy in 16 h study (F = 10.996,P〈0.01) and 8 Gy in 32 h study (F=17.019,P〈0.01), respectively. Time-response studies showed that MAP1LC3B mRNA increased and reached to peak at 16 h in 2 Gy dose-effect study (F = 16.284,P〈0.01) , 32 h in 8 Gy study (F =9.030,P 〈0.05) and 16 h in 12 Gy study (F=20.315, P 〈 0.05) , respectively. Dose-response studies of beelin 1 mRNA showed that beelin 1 increased after 2, 4 and 8 Gy irradiation. Time-response studies showed that beclin 1 increased in a time-dependent manner after 8 Gy irradiation. 16 h dose-effect study of beclin 1 protein showed the increase of beelin 1 expression, and 2 Gy time-response study showed that beelin 1 expression increased and reached the peak at 8 h. Conclusions X-ray irradiation could induce autophagy in MCF-7 cells, beelin 1 might play an important role in the process of autophagy.